Hepatitis C virus (HCV) is a single-stranded RNA virus in the Flaviviridae family of viruses. It is estimated that approximately 170 million people worldwide, and at least 4 million people in the United States, have been infected with HCV (Thomas D L, Astemborski J, Rai R M, Anania F A, Schaeffer M, Galai N, Nolt K, Nelson K E, Strathdee S A, Johnson L, Laeyendecker O, Boitnott J, Wilson L E, Vlahov D., The Natural History of Hepatitus C Virus Infection. JAMA 2000; 284 (4): 450-456). In the US, more people die of HCV than HIV infection (Ly, K., Xing J, Klevens R M, Jiles R B, Ward J W, Holmberg S D. The Increasing Burden of Mortality From Viral Hepatitis in the United States Between 1999 and 2007. Annals of Internal Medicine 156, 271-278 (2012).) Thus, infection with HCV represents a significant, worldwide health problem.
In most people, acute infection with HCV generally results in mild symptoms such as fatigue, decreased appetite, and flu-like symptoms. By convention, acute hepatitis refers to the presence of clinical signs or symptoms of hepatitis for a period of 6 months or fewer after the presumed time of exposure. In some instances, however, the newly infected individual remains asymptomatic. While some individuals can spontaneously clear the virus, approximately 85% of people infected with HCV will develop chronic hepatitis C, which is defined as persistent viremia occurring at least 6 months after initial exposure (Blackard J T, Shata M T, Shire N J, Sherman K E., Acute Hepatitus C Virus Infection: A Chronic Problem., Hepatology 2008; 47(1):321-331). Chronic infection with HCV is a leading cause of liver cancer and end-stage liver disease. It is also the most common reason for liver transplantation in the U.S. Currently, the standard treatment for HCV infections is pegylated interferon-α (IFN-α) combined with ribavirin. Successful treatment resolves chronic HCV infection, thereby markedly reducing HCV related morbidity and mortality, but the pegylated IFN-α/ribavirin regimen is effective in less than 45% of patients, is expensive and has many adverse effects. More recently, a triple therapy comprising pegylated-IFN-α, ribavirin, and an HCV protease inhibitor has been recommended. Although this new regimen should be more efficacious than treatment with pegylated-interferon-α/ribavirin, a sizeable proportion of patients may fail to respond and patients treated with this regimen will experience the adverse effects seen with pegylated-IFN-α/ribavirin therapy. Thus, a method of identifying patients who are unlikely to respond to treatment with interferon-based therapies is urgently desired so that these patients can be spared the expense and adverse effects associated with futile treatment. In addition, the failure of some patients to respond to treatment indicates the need for new treatments for hepatitis C infections.
Increasing evidence suggests that host genetic factors influence both the natural course of chronic HCV infection and response to therapy (Lauer G M, Walker B D. Hepatitis C virus infection. N Engl J Med 2001 Jul. 5; 345(1):41-52; Manns M P, McHutchison J G, Gordon S C, Rustgi V K, Shiffman M, Reindollar R, et al. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomised trial. Lancet 2001 Sep. 22; 358(9286):958-965; Fried M W, Shiffman M L, Reddy K R, Smith C, Marinos G, Goncales F L, Jr., et al. Peginterferon α-2a plus ribavirin for chronic hepatitis C virus infection. N Engl J Med 2002 Sep. 26; 347(13):975-982; Kau A, Vermehren J, Sarrazin C. Treatment predictors of a sustained virologic response in hepatitis B and C. J Hepatol 2008 October; 49(4):634-651). For example, in two cohorts of pregnant women infected under similar conditions from immunoglobulin preparations contaminated with a single strain of HCV, half spontaneously cleared the infection and half progressed to chronic hepatitis C (Grakoui A, Shoukry N H, Woollard D J, Han J H, Hanson H L, Ghrayeb J, et al. HCV persistence and immune evasion in the absence of memory T cell help. Science 2003 Oct. 24; 302(5645):659-662; Knapp S, Yee L J, Frodsham A J, Hennig B J, Hellier S, Zhang L, et al. Polymorphisms in interferon-induced genes and the outcome of hepatitis C virus infection: roles of MxA, OAS-1 and PKR. Genes Immun 2003 September; 4(6):411-419). Among chronically infected patients, response to treatment differs, even between cases with similar HCV-RNA levels and identical genotypes (Thio C L. Host genetic factors and antiviral immune responses to hepatitis C virus. Clin Liver Dis 2008 August; 12(3):713-26, xi.; Yee L J. Host genetic determinants in hepatitis C virus infection. Genes Immun 2004 June; 5(4):237-245; Muller R. The natural history of hepatitis C: clinical experiences. J Hepatol 1996; 24(2 Suppl):52-54). The response rates are strongly associated with ethnicity (Conjeevaram, H. S. et al. Peginterferon and ribavirin treatment in African American and Caucasian American patients with hepatitis C genotype 1. Gastroenterology, 131:470-7 (2006)). Previous reports revealed the influence of genetic polymorphisms of human leukocyte antigens (HLA) (Sheppard, P. et al. IL-28, IL-29 and their class II cytokine receptor IL-28R. Nat Immunol 4, 63-8 (2003); Robek, M. D., Boyd, B. S. & Chisari, F. V. Lambda interferon inhibits hepatitis B and C virus replication. J. Virol. 79, 3851-3854 (2005)), killer immunoglobulin-like receptors (KIRs) (Lauterbach, H. et al. Mouse CD8alpha+DCs and human BDCA3+DCs are major producers of IFN-lambda in response to Poly I:C. J Exp Med 207, 2703-17), cytokines (WO 00/08215), chemokines and interleukins as well as interferon-stimulated genes on HCV infection outcomes (Lasfar, A. et al. Characterization of the mouse IFN-lambda ligand-receptor system: IFN-lambdas exhibit antitumor activity against B16 melanoma. Cancer Res 66, 4468-77 (2006); Phillips, J. E. & Corces, V. G. CTCF: master weaver of the genome. Cell 137, 1194-211 (2009); Shyu, A. B., Wilkinson, M. F. & van Hoof, A. Messenger RNA regulation: to translate or to degrade. EMBO J. 27, 471-81 (2008); Conjeevaram, H. S. et al. Peginterferon and ribavirin treatment in African American and Caucasian American patients with hepatitis C genotype 1. Gastroenterology 131, 470-7 (2006); Ghany, M., Nelson, D. R., Strader, D. B., Thomas, D. L. & Seeff, L. B. An update on treatment of genotype 1 chronic hepatitis c virus infection: 2011 practice guidelines by the American association for the Study of Liver Diseases. Hepatology, December 12 (doi: 10.1002/hep.25524) (2011).
Previous studies have used a candidate gene approach based on a priori knowledge of the potential role of a gene in HCV infection. However, previous data do not allow accurate prediction of spontaneous clearance or response to treatment (Robek, M. D., Boyd, B. S. & Chisari, F. V. Lambda interferon inhibits hepatitis B and C virus replication. J. Virol. 79, 3851-3854 (2005)). In 2009, several groups reported results from independent genome-wide association studies (GWAS) that identified single nucleotide polymorphisms (SNPs) in the IFNL3 (IL28B) gene region that are associated with response to pegylated IFN-α/ribavirin treatment among patients with chronic hepatitis C, as well as spontaneous clearance of HCV infection. For example, U.S. Patent Publication No. 2011/0165124 by Bochud et al, which is incorporated herein in its entirety by reference, discloses numerous SNPs associated with both response to interferon-based treatment of HCV, and spontaneous clearance. Among the SNPs identified in these GWAS, the genotype based on rs12979860 is currently accepted as the best predictor of spontaneous clearance and treatment response (Rauch, A. et al. Genetic variation in IL28B Is associated with chronic hepatitis C and treatment failure: a genome-wide association study. Gastroenterology 138, 1338-1345 (2010); Thomas, D. L. et al. Genetic variation in IL28B and spontaneous clearance of hepatitis C virus. Nature 461, 798-801 (2009); Ge, D. et al. Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance. Nature 461, 399-401 (2009); Suppiah, V. et al. IL28B is associated with response to chronic hepatitis C interferon-alpha and ribavirin therapy. Nat Genet. 41, 1100-4 (2009); Tanaka, Y. et al. Genome-wide association of IL28B with response to pegylated interferon-alpha and ribavirin therapy for chronic hepatitis C. Nat Genet. 41, 1105-9 (2009)). A single nucleotide polymorphism (SNP) rs12979860 is located approximately 3 kb upstream of the IFNL3 (IL28B) translational start site. Commercial laboratory tests based on rs12979860 are now available for predicting a patient's probability of responding to treatment.
Compared to persons of European ancestry, African American patients have a higher frequency of chronic hepatitis C and a poorer response to therapy with IFN-α/ribavirin. Racial differences in the frequency of GWAS marker rs12979860 do not completely explain these disparities. Identification of a genetic marker that has optimal predictive values in all population groups would improve clinical decision models for treatment of chronic hepatitis C and help deliver personalized medicine to all HCV-infected patients.
While current tests have proved to be useful in identifying responders to treatment of chronic HCV infection, there remains a need for a more robust and accurate test for predicting spontaneous clearance and response to treatment. Moreover, current tests require the isolation and genotyping of nucleic acid molecules from an individual. Finally, as noted above, there remains a percentage of the population who do not respond to treatment for chronic HCV infection with current therapies. Thus, a need exists for novel methods and treatments for these patients. The present invention satisfies these needs and provides other benefits as well.